This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In addition to their functions as cellular redox cofactors, an ever-widening array of non-redox functions has been found for the pyridine nucleotide coenzymes in cellular regulation. First to be identified were bacterial toxins, including those responsible for the symptomology of cholera, diphtheria and pertussis. Evolutionarily related proteins include the multifunctional nuclear protein poly-ADPRibose synthase (PARP) involved in repair of DNA and apoptosis and ADPRibosyltransferases, both cytoplasmic and extracellular, that have been implicated in immunocompetency and immunoregulation. An evolutionarily distinct family of enzymes identified as CD38 correlates with cell transformation and cell differentiation and was found to be a dual functional enzyme. CD38 catalyzes the hydrolysis of NAD and cyclic-ADPribose (cADPR) as well as possesses ADP-Ribosyl cyclase activity. The product, cADPR, functions as a second messenger in calcium regulation while CD38 is critical for the innate immune response. A third distinct family of NAD -dependent enzymes is the histone deacetylase designated as SIR2. It catalyzes the cleavage of the nicotinamide-ribosyl bond with concomitant transfer of the acetyl group to generate a mixture of O2'and O3'-Acetyl-ADPRibose. Mutations in SIR2 in yeast and Drosophila have led to increases in life span of up to two fold. A fourth family of NAD -dependent enzymes is the tRNA phosphotransferase, TPT1, that is the final step in maturation of tRNA. TPT1 catalyses the cleavage of the nicotinamide-ribosyl bond and then transfers the terminal 2'-phosphate of tRNA to generate ADPRibose-1'-2'-cyclicphosphate. Our lab is collaborating in mechanistic studies of the enzymatic functions as well as exploring the design of inhibitors and alternative substrates that can be used to probe their biological functions as well as their potential for therapeutic intervention. There are structural data for all four families hence the facilities of the Computer Graphics Laboratory play a central role in these investigations.